Early molecular events associated with nitrogen deficiency in rice seedling roots.

Nitrogen (N) deficiency is one of the most common problems in rice. The symptoms of N deficiency are well documented, but the underlying molecular mechanisms are largely unknown in rice. Here, we studied the early molecular events associated with N starvation (-N, 1 h), focusing on amino acid analysis and identification of -N-regulated genes in rice roots. Interestingly, levels of glutamine rapidly decreased within 15 min of -N treatment, indicating that part of the N-deficient signals could be mediated by glutamine. Transcriptome analysis revealed that genes involved in metabolism, plant hormone signal transduction (e.g. abscisic acid, auxin, and jasmonate), transporter activity, and oxidative stress responses were rapidly regulated by -N. Some of the -N-regulated genes encode transcription factors, protein kinases and protein phosphatases, which may be involved in the regulation of early -N responses in rice roots. Previously, we used similar approaches to identify glutamine-, glutamate-, and ammonium nitrate-responsive genes. Comparisons of the genes induced by different forms of N with the -N-regulated genes identified here have provided a catalog of potential N regulatory genes for further dissection of the N signaling pathwys in rice.

Identification of early ammonium nitrate-responsive genes in rice roots.

Ammonium has long been used as the predominant form of nitrogen source for paddy rice (Oryza sativa). Recently, increasing evidence suggests that nitrate also plays an important role for nitrogen acquisition in the rhizosphere of waterlogged paddy rice. Ammonium and nitrate have a synergistic effect on promoting rice growth. However, the molecular responses induced by simultaneous treatment with ammonium and nitrate have been less studied in rice. Here, we performed transcriptome analysis to identify genes that are rapidly regulated by ammonium nitrate (1.43 mM, 30 min) in rice roots. The combination of ammonium and nitrate preferentially induced the expression of nitrate-responsive genes. Gene ontology enrichment analysis revealed that the early ammonium nitrate-responsive genes were enriched in "regulation of transcription, DNA-dependent" and "protein amino acid phosphorylation" indicating that some of the genes identified in this study may play an important role in nitrogen sensing and signaling. Several defense/stress-responsive genes, including some encoding transcription factors and mitogen-activated protein kinase kinase kinases, were also rapidly induced by ammonium nitrate. These results suggest that nitrogen metabolism, signaling, and defense/stress responses are interconnected. Some of the genes identified here may be involved in the interaction of nitrogen signaling and defense/stress-response pathways in plants.

Acute multiple focal neuropathies and delayed postanoxic encephalopathy after alcohol intoxication.

Acute-onset alcohol-associated neuropathy is only occasionally reported, and delayed postanoxic encephalopathy is rare. Here, we report a male who developed acute multiple focal neuropathies and later delayed postanoxic encephalopathy after alcohol intoxication. He had hypoxia and rhabdomyolysis, presenting with acute renal failure initially, and cardiopulmonary support, including mechanical ventilation, led to improvement of the patient at the acute stage. He suffered from bilateral hand numbness and mild weakness of the right lower limb thereafter. Nerve-conduction study revealed no pickup of compound muscle action potential or sensory nerve action potential in the bilateral ulnar nerve, but showed attenuated amplitude of compound muscle action potential in the right femoral nerve. Multiple focal neuropathies were suspected, and he received outpatient rehabilitation after being discharged. However, the patient developed gradual onset of weakness in four limbs and cognitive impairment 23 days after the hypoxia event. Brain computed tomography showed low attenuation over bilateral globus pallidus, and brain magnetic resonance imaging disclosed diffuse increased signal intensity on T 2-weighted images and fluid-attenuated inversion recovery in bilateral white matter. He was admitted again under the impression of delayed postanoxic brain injury. Supportive treatment and active rehabilitation were given. He had gradual improvement in motor and functional status after rehabilitation. He could walk with festinating gait under supervision, and needed only minimal assistance in performing activities of daily living approximately 1 year later.

Application of an inducible transposon with anther culture in generation of di-haploid homologous mutants.

BACKGROUND: Insertional mutagenesis represents one of the most effective ways to acquire information about a plant gene's function. However, it is hindered by the autosomal genome being diploid and therefore, most mutations being recessive. The problem is addressed by inducing the transposition during anther culture so that selected mutations can be transmitted and then regenerated to a homozygous state. RESULTS: To this end, we treated transgenic rice floral tissues containing the inducible transposon with an inducer, salicylic acid. Excision events were detected in regenerated calli and subsequent plantlets. DNA blot and PCR assay were used to determine the homogeneity of knockout mutants. About 5% of the mutants containing transposition events were homozygous. Furthermore, the inducible transposon was active during calli regeneration. CONCLUSIONS: This strategy could be applicable to improve transposition efficiency in microspore development stages to create stable di-haploid mutants in plants.

Ds transposon is biased towards providing splice donor sites for exonization in transgenic tobacco.

Insertion of transposed elements into introns can lead to their activation as alternatively spliced cassette exons, an event called exonization, which can enrich the complexity of transcriptomes and proteomes. In this study, the first exonization event was detected when the modified rice EPSPS marker gene was inserted with the Ac transposon 5' end, which provided a splice donor site to yield abundant novel transcripts. To assess the contribution of splice donor and acceptor sites of transposon sequences, we inserted a Ds element into each intron of the EPSPS marker gene. This process yielded 14 constructs, with the Ds transposon inserted in the forward and reverse direction in each of the 7 introns of the EPSPS marker gene. The constructs were transformed into tobacco plants, and novel transcripts were identified by RT-PCR with specific primers. Exonization of Ds in EPSPS was biased towards providing splice donor sites of the inserted Ds sequence. Additionally, when the Ds inserted in reverse direction, a continuous splice donor consensus region was determined by offering 4 donor sites in the same intron. Information on these exonization events may help enhance gene divergence and functional genomic studies.